International Journal of Molecular Sciences Article Cytoplasmic Inter-Subunit Interface Controls Use-Dependence of Thermal Activation of TRPV3 Channel Lucie Macikova 1,2 , Lenka Vyklicka 1, Ivan Barvik 3, Alexander I. Sobolevsky 4 and Viktorie Vlachova 1,* 1 Department of Cellular Neurophysiology, Institute of Physiology Czech Academy of Sciences, 142 20 Prague, Czech Republic 2 Department of Physiology, Faculty of Science, Charles University, 128 00 Prague, Czech Republic 3 Division of Biomolecular Physics, Institute of Physics, Faculty of Mathematics and Physics, Charles University, 121 16 Prague, Czech Republic 4 Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032, USA * Correspondence:
[email protected]; Tel.: +420-296-442-711 Received: 23 July 2019; Accepted: 13 August 2019; Published: 16 August 2019 Abstract: The vanilloid transient receptor potential channel TRPV3 is a putative molecular thermosensor widely considered to be involved in cutaneous sensation, skin homeostasis, nociception, and pruritus. Repeated stimulation of TRPV3 by high temperatures above 50 ◦C progressively increases its responses and shifts the activation threshold to physiological temperatures. This use-dependence does not occur in the related heat-sensitive TRPV1 channel in which responses decrease, and the activation threshold is retained above 40 ◦C during activations. By combining structure-based mutagenesis, electrophysiology, and molecular modeling, we showed that chimeric replacement of the residues from the TRPV3 cytoplasmic inter-subunit interface (N251–E257) with the homologous residues of TRPV1 resulted in channels that, similarly to TRPV1, exhibited a lowered thermal threshold, were sensitized, and failed to close completely after intense stimulation. Crosslinking of this interface by the engineered disulfide bridge between substituted cysteines F259C and V385C (or, to a lesser extent, Y382C) locked the channel in an open state.